HIV pathogenesis is characterized by the progressive loss of CD4+ T cells leading to a state of extreme immunodeficiency. Rather than simply due to direct infection and death of target CD4+ T cells, this is now appreciated to be more extensively due to overt and sustained chronic immune activation. Levels of immune activation can vary significantly between individuals, and this represents a major contributor to the differential kinetics of CD4+ T cell decline observed among infected individuals. We have previously identified that the viral replication capacity (vRC) of the transmitted virus is a significant predictor of differential immune activation observed during acute HIV infection (Claiborne et al., PNAS, 2015). Specifically, the vRC of the transmitted virus is positively correlated with the level of inflammatory cytokines and the frequency of activated T cells and is a significant predictor of longitudinal CD4+ T cell decline independent of viral load or protective HLA class I alleles. However, despite extensive data linking vRC to immune activation, and immune activation to CD4+ T cell loss, the precise mediator of this immune activation has not yet been elucidated. This is largely due to limitations in the ability to sample plasma and mucosal tissues during the eclipse phase of the infection in human cohorts, and the confounding effects of baseline immune activation and co-infections on these early events. As such, these studies would benefit from a small animal model of HIV infection that would accurately recapitulate the salient aspects of immune activation and CD4+ T cell loss. Here, we propose to use the bone marrow-liver-thymus (BLT) humanized mouse model to elucidate the mechanism(s) underlying HIV-associated bystander T cell activation and death linked to chronic immune activation. Our preliminary data demonstrates that the BLT mouse model recapitulates vRC-associated differential immune activation and CD4+ T cell loss, in that mice infected with high vRC viruses displayed significantly higher frequencies of CD38+/HLA-DR+ and PD-1+ CD8+ T cells and significantly increased CD4+ T cell decline. Moreover, congruent with human studies, our preliminary data in BLT mice suggests a role for the innate immune system in mediating this acute immune activation. Therefore, we intend to use the BLT mouse model of HIV infection to interrogate the extent to which immune activation and CD4+ T cell loss are driven by interactions between vRC and the innate immune system. To this end, we propose to infect BLT mice with an array of viruses exhibiting significant differences in vRC and to monitor the levels of inflammatory cytokines as well as peripheral and mucosal cellular immune activation. Additionally, we propose to employ RNA-seq analysis to describe gene pathways differentially expressed between BLT mice infected with low and high vRC viruses. In concert, we will evaluate the contributions of the innate immune system by comparing myeloid subset reconstitution levels with inflammatory gene expression. Collectively, this analysis will allow us to identify candidate gene pathways responsible for increased innate immune activation and exacerbated CD4+ T cell loss. Finally, we will use this information to identify novel targets and to test therapeutic interventions designed to blunt early immune activation and the resultant pathogenesis.